OP0309 WES ANALYSIS OF RARE FAMILIES POINTS TO A PATHOGENIC ROLE FOR THE cGAS/STING-TYPE I IFN AXIS IN SSc
Authors: P. Maus, V. Smith, T. Du Four, M. Vanthuyne, B. Lauwerys, N. Limaye
Journal: Annals of the Rheumatic Diseases (2022)

Background Systemic sclerosis (SSc) is a rare (prevalence ±20/100,000), chronic condition characterized by vascular, immunological and connective tissue abnormalities. Genetic factors contribute to disease, as evidenced by a 15-fold increased relative risk in relatives of patients. Case-control studies have identified numerous associated loci; the identification and functional validation of risk alleles has however been elusive. No monogenic or oligogenic causes of SSc have been demonstrated, likely due to the infrequency of familial clustering (≤ 1.6% of cases) in this disease. We are fortunate to have access to DNA samples from five such families, with two affected first-degree relatives each: To our knowledge the only series of its kind. Objectives The goal of our research is to identify and functionally validate genes that drive SSc pathogenesis in these families, thereby shedding light on fundamental disease mechanisms. In light of the apparent lack of a multi-generational effect, we hypothesize that SSc may be transmitted as a di/oligogenic trait in these families, with two-to-a-few genes acting together to cause disease. Methods Whole exome sequencing (WES) was performed on all available blood-DNA (Figure 1). We filtered for variants that are (a) shared by both affected individuals within each family, (b) absent-to-rare in the general population, and (c) predicted to affect protein function by multiple in silico tools. Candidate gene prioritization and pathway analysis is followed by in vitro testing of a subset of SSc-variants, to assess for their effects on protein function. Where two strong candidate genes are identified in the same family, in vitro validation of both variants will be followed by the generation of single- vs. double- knock-in mice, to test for di/oligogenic effects of variant combinations on SSc-related phenotypes. Figure 1. SSc families. Filled symbols: affected. Arrows: WES on blood-DNA. lSSc: Limited SSc; lcSSc: Limited cutaneous SSc; dcSSC: Diffuse cutaneous SSc. cGAS/STING (red) and IFN I (black) - related candidate genes. Results WES yielded 23-45 genes per family with variants that satisfied the above criteria. Prioritization of candidates was based on systematic annotation of all 180 genes for expression pattern, function, and potential impact of the SSc-variants. Our data suggest familial SSc is genetically heterogeneous, with little sharing of candidates between families. We therefore assessed for pathways represented in gene-lists from multiple families, and found that participants in the cGAS/STING - type I-IFN axis are identified in all (Figure 1). This would suggest a unifying disease mechanism. cGAS/STING is a sentinel system responsible for the production of anti-viral type I interferons (IFN-α/β) and pro-inflammatory cytokines in the presence of cytoplasmic double stranded DNA (dsDNA). cGAS/STING-dependent self-dsDNA sensing has been implicated in silica-induced inflammation: a risk factor in SSc and pulmonary fibrosis. A type I IFN-response signature is identified in SSc patients and correlates with disease severity. Mutations in two of our candidates cause Mendelian diseases with phenotypic overlaps with SSc, and all but one cause disease-relevant phenotypes when perturbed in mice. Finally, preliminary in vitro data suggest gain-of-function effects of the variant identified in TMEM173/ STING, and loss-of-function effects in NT5E /CD73 and PRKD2 (the last with two different variants identified in two families; Figure 1). All would lead to increased IFN-I signaling. Conclusion We report, for the first time, WES analysis of a familial SSc series of unprecedented size for this rare disease. The identification of potential “driver” gene variants in cGAS/STING-type I IFN signaling regulators across all families points to an important pathogenic role for this pathway in SSc. Disclosure of Interests Pierre Maus: None declared, Vanessa Smith: None declared, Tessa Du Four: None declared, Marie Vanthuyne: None declared, Bernard Lauwerys Shareholder of: UCB Pharma, Employee of: Currently an employee of UCB Pharma, Nisha Limaye: None declared